CN106916901B - Wax-apple EST-SSR molecular labeling - Google Patents
Wax-apple EST-SSR molecular labeling Download PDFInfo
- Publication number
- CN106916901B CN106916901B CN201710318661.2A CN201710318661A CN106916901B CN 106916901 B CN106916901 B CN 106916901B CN 201710318661 A CN201710318661 A CN 201710318661A CN 106916901 B CN106916901 B CN 106916901B
- Authority
- CN
- China
- Prior art keywords
- wax
- apple
- ssr
- dna
- artificial sequence
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6888—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms
- C12Q1/6895—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for detection or identification of organisms for plants, fungi or algae
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Analytical Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Organic Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Health & Medical Sciences (AREA)
- Biotechnology (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Immunology (AREA)
- Mycology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Botany (AREA)
- Biophysics (AREA)
- Physics & Mathematics (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
Abstract
The present invention provides wax-apple EST-SSR molecular labelings, belong to field of molecular marker, the site SSR of upper frequency can be obtained by carrying out SSR marker exploitation using wax-apple transcript profile data, 22 pairs of primers have been determined, and type is abundant, provides more abundant reliable label selection for wax-apple analysis of genetic diversity and genetic map construction.
Description
Technical field
The invention belongs to field of molecular marker, and in particular to wax-apple EST-SSR molecular labeling.
Background technique
Wax-apple [Syzygium samarangense(Blume) Merri.et Perry] originate in the Malay Peninsula and peace
Dammam archipelago, be Myrtaceae (Myrtaceae) Syzygium (Syzygium) tropical evergreen fruit tree, alias wax jambo, Nan Yang
Portugal, Java Portugal, sweet tea mist, mirabilitum crystallina pomegranate etc..Wax-apple is rich in a variety of medicinal ingredients, such as flavonoids, chalcone, hexane, tool
There are antidiarrheal, analgesia, antibacterial, antihyperglycemic, anticholinesterase, anti-inflammatory, raising immunity and other effects.Wax-apple fruit colour is gorgeous,
Form is unique, and fresh and sweet tasty and refreshing, mouthfeel is good, is rich in phenolic substances, is one of the fruit of great health value, by domestic and international consumer
Like.There is extensive plantation on Malaysia, Thailand, Indonesia and other places, is the important component of local fruit industry, fresh fruit
Past American-European countries in constant demand.The area of China's early start commercialization plantation wax-apple is Taiwan, in recent years, with traditional hot-zone advantage
The falling of the prices such as fruit tree such as lichee, longan, citrus, wax-apple become emerging tropical fruit tree, and cultivated area gradually expands, main product
Area is the ground such as Taiwan, Hainan, Fujian, Guangdong and Guangxi, according to incompletely statistics, China's wax-apple cultivated area 6.87 ten thousand in 2014
Mu, 13.6 ten thousand tons of yield, 30 ~ 50 yuan of per kilogram price, high financial profit has development potentiality.
Wax-apple has with genetic diversity abundant, but correlative study report is fewer.Taiwan is according to fruit color by lotus
Mist is divided into big (depth) Red Indian race, light red kind, pink kind, five classes (positive loyalty of woods etc., 2004) such as blueness kind and white race.Wang Jiabao etc.
(2004) Studies on Isozymes is carried out to 11 parts of wax-apple resources in Hainan, the results showed that 11 parts of resources are polymerized to red according to color difference
Color pericarp, white pericarp, 3 major class of green pericarp, red peel are close with the white pericarp more green pericarp of affiliation.But what bridge etc.
(2006) ISSR analysis carried out to 12 parts of wax-apple resources and 2 wax-apple sibling species, cluster analysis result show wax-apple not with
Fruit colour is identical to be gathered for one kind, and with the maturity period it is close gather for one kind, therefore he points out that the maturity period may close in wax-apple relationship
It plays an important role in system and classification position.Wax-apple is planted after the states such as Java, Malaysia introduce China in long-term adaptability
Many local varieties and resource are produced under training, these local varieties are mostly named with fruit color, such as ' bright red wax-apple ', ' pink
Wax-apple ', ' pake purpke mist ' etc., this is very likely to cause homonym and synonym, not only collects, protects to Wax Apple Germplasm
It deposits, identify and using difficulty is caused, also constrain the breeding and popularization and application of excellent variety.
DNA molecular marker is the important means for identifying Genetic Diversity of Germplasm, wherein microsatellite sequence (i.e. simple weight
Complex sequences Simple sequence repeat, SSR) molecular labeling is because of its site-specific, high polymorphism, high stability, repetition
Property it is good, be widely used in the advantages that codominance analysis of genetic diversity, cultivar identification, genetic map establish etc..
EST-SSR label derives from the transcriptional domain of DNA sequence dna, more than the SSR marker cross-species amplification developed based on genome sequence
Height, also more economical convenience are widely used in recent years in the SSR molecular marker exploitation of various plants, Rosa roxburghii Tratt, onion,
It is had been reported that on the gardening plants such as Lonicera edulis, sponge gourd.The present invention carries out SSR marker using the data that transcript profile sequencing obtains
Search analyzes its distribution and composition characteristic, carries out preliminary usability evaluation, more to cultivar identification, the germ plasm resource for wax-apple
Sample analysis establishes Core Germplasms and molecular mark lays the foundation.
Summary of the invention
The purpose of the present invention is to provide wax-apple EST-SSR molecular labelings, carry out SSR marker using wax-apple transcript profile data
Exploitation can obtain the site SSR of upper frequency, and type is abundant, provides for wax-apple analysis of genetic diversity and genetic map construction
More abundant reliable label selection.
To achieve the above object, the present invention adopts the following technical scheme:
Wax-apple EST-SSR molecular labeling number is WASSR3, WASSR12-13, WASSR37-39, WASSR41-42,
WASSR44、WASSR50、WASSR55、WASSR60、WASSR63、WASSR70、WASSR72、WASSR74、WASSR80、
WASSR85, WASSR87-88, WASSR91-92, the primer sequence of the corresponding wax-apple EST-SSR molecular labeling of each number is such as
Shown in SEQ ID NO.1-44.
Application of the wax-apple EST-SSR molecular labeling in wax-apple analysis of genetic diversity and genetic map construction.
Specific method of the invention is:
(1) wax-apple genomic DNA is extracted using CTAB method；
(2) search of the site SSR is carried out using MISA program (http://pgrc.ikp-gatersleben.de/misa),
It is respectively to be scanned for for standard for 6,5,5,5 and 5 times with two to the minimum number of repetition of Hexanucleotide.With 3.0 software of Primer
Design of primers and evaluation are carried out to the sequence before and after SSR repetitive unit, every SSR generates 3 primers.Primer sequence length 18 ~
27bp, G/C content 40% ~ 60%, 57 ~ 63 DEG C of annealing temperature, the Tm value of upstream and downstream primer differs≤2 DEG C, it is contemplated that amplified production is long
Spend 100 ~ 280 bp；And without secondary structure and dimer；
(3) EST-SSR primer screening: PCR reaction system is 20ul, wherein 4 μ l, 5U Taq enzyme of 2.5mmol/L dNTP
0.3 μ l, 100 ng DNA 1.5 μ l, the upstream and downstream primer of 10 μm of ol/L each 1 μ l, 10 × Buffer(Mg2+) 2.5 μ l,
Add ddH2O is mended to 20 μ l.PCR amplification program are as follows: 95 DEG C of 5 min of initial denaturation；Then 35 circulations are carried out, each circulation includes
94 DEG C of 30 s of denaturation, 56 DEG C of 30 s(annealing temperatures of annealing are different because of different primers), 72 DEG C of 1 min of extension；Last 72 DEG C of extensions
7 min。
The present invention has the advantages that
The present invention is to 21 153 Unigene(4.75Mb from 3 wax-apple kind fruits) sequence carry out EST-SSR search
Rope, obtains 11 641 sites SSR, the frequency of occurrences 55.03%, be higher than Lonicera edulis (32.51%), Rosa roxburghii Tratt (20.37%),
The orchard fruits such as lichee (16.35%), citrus (21.74%, 5.2kb), pears (7.1%, 7.2%), also above radish (23.79%), silk
The herbaceous plant such as melon (14.97%), capsicum (7.84%), lavender (9.88%).From the point of view of forefathers are to the result of study of plant, greatly
The EST-SSR of most plants is based on two, Trinucleotide repeats type, and leading repetition motif is different because of species.Present invention hair
Show wax-apple based on mononucleotide repeat type (43.44%), followed by dinucleotides repeat type (30.98%) and trinucleotide
Repeat type (24.38%), though four, five and Hexanucleotide repeat type occur, proportion very little (1.21%), this and cottonrose hibiscus
Lee Rong, Cortex Eucommiae are similar with the result of study of Fructus Forsythiae, are different from lichee, loquat and Lonicera edulis.The mononucleotide of wax-apple repeats
The abundantest with A/T in motif, the fruit tree research result such as this is with citrus and Fu-Rong plums is consistent；Dinucleotides repeats AG/ in motif
CT is most, identical as the most fruit trees having been reported, and is different from lichee (GA/TC)；Trinucleotide repeats motif is with CCG/CGG
It is main, it is different based on AAG/CTT from Kiwi berry, citrus and strawberry etc., also different from lichee (GAA/TTC) and pears (ACC/GGT).
The present invention selects 100 pairs of SSR primer of 14 ~ 25bp to synthesize from 25734 pairs of primer kinds at random, wherein 67 pairs
Primer can amplify ideal PCR product, and effective amplification rate is 67.00%.Wherein, 22 pairs of SSR primers presentations are polymorphic in 30 pairs of primers
Property, occupy the 73.33% of effect primer.This ratio is higher than Rosa roxburghii Tratt (52.17%), Fu-Rong plums (42.5%), Lonicera edulis
(62.5%), a variety of fruit trees such as lichee (66.67%).This illustrates that the site SSR in wax-apple est sequence is more, primer amplification efficiency
It is higher, and its polymorphism is also relatively high.22 pairs of polymorphic differences primer pairs, 30 parts of wax-apple materials carry out UPGMA cluster, are divided
It is threshold value with genetic distance 0.68 for 2 major class, the 1st class can be divided into 4 groups, gathers so that fruit colour or source place are close for one kind,
2nd class can be divided into 2 groups, distinguished with fruit colour difference, more accurately reflect the difference between 30 parts of wax-apple materials.
The present invention is higher using the SSR marker availability that wax-apple transcript profile data mining goes out, and grinds for Wax Apple Germplasm genetic diversity
Study carefully, genetic map construction, gene mapping and cloning and molecular mark etc. are laid a good foundation.
Detailed description of the invention
Polymorphism of Fig. 1 primer 63 in 30 wax-apple materials.Kind number title is shown in Table 1.
For Fig. 2 for trying the UPGMA dendrogram of wax-apple material, kind number title is shown in Table 1.
Specific embodiment
1 materials and methods
1.1 transcript profile data sources
Wax-apple transcript profile data source carries out Illumina high throughput deep sequencing to fruit in 2016 in this seminar
As a result.Acquire that ' purplish red ', ' black pearl ', ' blueness bores ' 3 wax-apple kind fruit, and each kind fetches from 3 differences when sequencing
The wax-apple fruit of single plant each 20, liquid nitrogen flash freezer, -80 DEG C of preservations.When sequencing, each single plant constructs 1 library.Sequencing commission north
Jing Baimaike Biotechnology Co., Ltd carries out RNA-Seq transcription using 2500 PE125 system of Illumina HiSeqTM
Group sequencing (no ginseng) carries out sequence assembling using Trinity, the valid data of 7.80G is obtained through filtering.
1.2 materials and its DNA are extracted
It is Inst. of Fruit Trees, Fujian Prov. Academy of Agricultural Sciences's wax-apple germplasm for the material of SSR primer screening and usability evaluation
30 parts of germ plasm resources (table 1) of resource garden.Extracting genome DNA is carried out using CTAB method.
1 wax-apple SSR polymorphism analysis material of table
1.3 sites transcript profile SSR identify and SSR design of primers
The search of the site SSR is carried out using MISA program (http://pgrc.ikp-gatersleben.de/misa), with two
It is respectively to be scanned for for standard for 6,5,5,5 and 5 times to the minimum number of repetition of Hexanucleotide.With 3.0 software of Primer to SSR
Sequence before and after repetitive unit carries out design of primers and evaluation, and every SSR generates 3 primers.Primer sequence 18 ~ 27bp of length,
G/C content 40% ~ 60%, 57 ~ 63 DEG C of annealing temperature, the Tm value of upstream and downstream primer differs≤2 DEG C, it is contemplated that and amplified production length 100 ~
280 bp；And without secondary structure and dimer.
1.4 EST-SSR primer screenings
PCR reaction system is 20ul, wherein 0.3 μ l, 100 ng DNA of 4 μ l, 5U Taq enzyme of 2.5mmol/L dNTP
1.5 μ l, the upstream and downstream primer of 10 μm of ol/L each 1 μ l, 10 × Buffer(Mg2+) 2.5 μ l, add ddH2O is mended to 20 μ l.
PCR amplification program are as follows: 95 DEG C of 5 min of initial denaturation；Then 35 circulations are carried out, each circulation includes 94 DEG C of denaturation 30 s, and 56 DEG C
30 s(annealing temperatures of annealing are different because of different primers), 72 DEG C of 1 min of extension；7 min of last 72 DEG C of extensions.Pcr amplification product
With 2% agar sugar detection, 6% denaturing polyacrylamide gel is detected, 160V voltage, 3h, is observed and is taken pictures after silver staining colour developing.
1.5 data statistics
The Unigene quantity and total Unigene ratio of number that SSR occurrence frequency is SSR, the frequency of occurrences of SSR are SSR's
The quantity ratio of number and total Unigene, SSR be evenly distributed distance be 1kb or more Unigene base number and SSR quantity it
Than.Using the method manually read tape, the clear band repeated on electrophoretogram is denoted as " 1 ", same position is without band or is not easy point
The weak band distinguished is denoted as " 0 ", establishes raw data matrix.Cluster drawing is carried out by Hierarchical Clustering using software NTsys2.10.
2 results and analysis
The distribution and design feature of SSR in 2.1 transcript profiles
87 538 Unigene(sequences are obtained after wax-apple transcript profile is assembled altogether and are always about 75 156 717 bp), it uses
For MISA software to Unigene(21 153 of 1kb or more, sequence is 47 491 726 bp) it scans for, discovery is wherein
Contain 11 641 sites SSR in 8 115 Unigene sequences, wherein 2 773 Unigene contain two or more
The site EST-SSR.SSR occurrence frequency is 38.36%, the frequency of occurrences 55.03%, and 1 SSR occurs in average 4.08kb, wherein
Composite S SR has 1432, accounts for 12.3%.6 kinds of SSR repeat type have, i.e., mononucleotide to Hexanucleotide repeats.Wherein monokaryon
Thuja acid, dinucleotides and the Trinucleotide repeats frequency of occurrences are dominant, respectively 43.44%, 30.98% and the 24.38% of the total SSR of Zhan；
Tetranucleotide, pentanucleotide and Hexanucleotide repeat type negligible amounts account for 0.92%, the 0.14% and 0.15%(table of sum respectively
2).
The number of repetition of wax-apple transcript profile SSR repetitive unit is distributed between 5 ~ 23 times, wherein 5 ~ 10 SSR are shared
8753, account for the 75.19% of sum；Secondly it is 11 ~ 20 SSR, shares 2880, accounts for the 24.74% of sum；20 repetitions with
On only 8, only account for 0.07%.The length of wax-apple transcript profile SSR is concentrated mainly on 10 ~ 256bp, and wherein length is less than 12bp
SSR have 2873 (24.68%), length 12 ~ 20bp SSR up to 7094, account for sum 60.94%, length 20bp with
On SSR have 920, only account for 7.90%.
Type, quantity and the distribution frequency of 2 wax-apple SSR of table
SSR motif repeat type and frequecy characteristic in 2.2 transcript profiles
11 641 sites SSR contain 72 kinds of repetition motifs altogether in wax-apple transcript profile, and mononucleotide to Hexanucleotide repeats to divide
There are not 2,4,10,24,15 and 17 kind.From the point of view of distribution frequency (table 3), occur at most with mononucleotide repeat type A/T, Zhan is total
The 40.15% of SSR accounts for mononucleotide repeats the sum of motif 92.43%.Followed by dinucleotides repeat type AG/CT, Zhan are total
The 26.99% of SSR accounts for the 87.13% of dinucleotides sum.In addition, with CCG/CGG, AGG/CCT in Trinucleotide repeats motif
With AAG/CTT dominance, 33.62%, 20.05% and the 16.98% of trinucleotide sum is accounted for respectively；Tetranucleotide, pentanucleotide and
Hexanucleotide repetition motif distribution is more dispersed, and the frequency of appearance is lower.
Different microsatellites repeat the frequency that motif (motif) occurs in the transcription of 3 wax-apple of table
2.3 wax-apple transcript profile SSR design of primers and screening
Design of primers is carried out using 8 115 Unigene sequences of the Primer3.0 to the site containing SSR, designs primer altogether
25 734 pairs.The primer pair wax-apple ' pink kind ' of 100 pairs of different repeat units (two, three, four, five, Hexanucleotide) is selected at random
DNA carries out SSR-PCR amplification to verify its validity.The result shows that 76 pairs of primers realize effectively amplification, 100 pairs of SSR primers are accounted for
76%.In 76 pairs of effective amplimers, 67 are consistent to (88.12%) pcr amplification product with expected size, there is 9 pairs
(13.43%) amplified production length is more than to be expected.
2.4 polymorphism analysis
30 parts of wax-apples are chosen, amplification is carried out using 30 pairs of effective EST-SSR primers and polymorphism is evaluated.Wherein draw for 22 pairs
There are polymorphic differences (tables 4) for object, occupy the 73.33% of effect amplimer.The polymorphic bands number that each pair of primer generates is 2 ~ 6
Between, 63 band are obtained in 23 pairs of primers, wherein polymorphic bands 52, and each pair of primer averagely generates 2.36 polymorphism pieces
Section, Fig. 1 are the amplification situation of primer 63.
4 22 pairs of wax-apple SSR primer information of table
Clustering is carried out using 22 pairs of polymorphism SSR primer pairs, 30 parts of wax-apple materials, at genetic distance 0.60, for examination
Material is divided into 2 major class (Fig. 2).1st class includes 26 parts of materials, is threshold value with genetic distance 0.68, such is divided into 4 groups,
Wherein with ' Indonesia great Ye ', ' black diamond ' and ' purplish red ' be the red peel wax-apple of representative gather be it is a kind of, with ' emerald ', ' Thailand
Green kind ' and ' Peth Jinda ' is that the green pericarp wax-apple of representative is got together, ' imperial Wen Shisheng ' and ' Indian red ' are classified as one kind,
Gather from ' green kind of the Changtai ' in same orchard, ' blueness is bored ' and ' pink kind of Changtai ' for one kind.2nd class includes 4 parts of materials, wherein
Red peel wax-apple (' ruby ', ' perfume ', ' agriculture section 4 ') is classified as one kind, and ' white lotus mist ' pericarp is white, is constituted a class by itself.
The foregoing is merely presently preferred embodiments of the present invention, all equivalent changes done according to scope of the present invention patent with
Modification, is all covered by the present invention.
SEQUENCE LISTING
<110>Inst. of Fruit Trees, Fujian Prov. Academy of Agricultural Sciences
<120>wax-apple EST-SSR molecular labeling
<130> 44
<160> 44
<170> PatentIn version 3.3
<210> 1
<211> 20
<212> DNA
<213>artificial sequence
<400> 1
tgtttcaact cgcacgactc 20
<210> 2
<211> 20
<212> DNA
<213>artificial sequence
<400> 2
aaacggtttt cattgatggc 20
<210> 3
<211> 20
<212> DNA
<213>artificial sequence
<400> 3
gtgtcatcga aaccctccaa 20
<210> 4
<211> 20
<212> DNA
<213>artificial sequence
<400> 4
ttgaatcttg caagcgactg 20
<210> 5
<211> 20
<212> DNA
<213>artificial sequence
<400> 5
gtcatgcttg ctctgtccaa 20
<210> 6
<211> 20
<212> DNA
<213>artificial sequence
<400> 6
ttctggattg caggaacctt 20
<210> 7
<211> 20
<212> DNA
<213>artificial sequence
<400> 7
gatggctggt gactcaggtt 20
<210> 8
<211> 20
<212> DNA
<213>artificial sequence
<400> 8
gaccctttcc ttgagtggaa 20
<210> 9
<211> 23
<212> DNA
<213>artificial sequence
<400> 9
ggaaaaggag agaattttct tga 23
<210> 10
<211> 20
<212> DNA
<213>artificial sequence
<400> 10
ttttccccca tgagagtgag 20
<210> 11
<211> 20
<212> DNA
<213>artificial sequence
<400> 11
gggttttcag ccaaaaattg 20
<210> 12
<211> 20
<212> DNA
<213>artificial sequence
<400> 12
cgagcgagag agagagatgg 20
<210> 13
<211> 20
<212> DNA
<213>artificial sequence
<400> 13
gagcttgttt atcgaaggcg 20
<210> 14
<211> 20
<212> DNA
<213>artificial sequence
<400> 14
aaactggcga caaaaacgag 20
<210> 15
<211> 20
<212> DNA
<213>artificial sequence
<400> 15
gtcgcgtcac cattatttca 20
<210> 16
<211> 20
<212> DNA
<213>artificial sequence
<400> 16
ggtttagtta agggcggctc 20
<210> 17
<211> 20
<212> DNA
<213>artificial sequence
<400> 17
gaggagtcgc tgaactgtcc 20
<210> 18
<211> 20
<212> DNA
<213>artificial sequence
<400> 18
cgcacccact gttttaggat 20
<210> 19
<211> 20
<212> DNA
<213>artificial sequence
<400> 19
aagccgaaga tgaagacgaa 20
<210> 20
<211> 20
<212> DNA
<213>artificial sequence
<400> 20
ggacatatat ccgcggtttg 20
<210> 21
<211> 20
<212> DNA
<213>artificial sequence
<400> 21
atatgaatcc cgggccttac 20
<210> 22
<211> 20
<212> DNA
<213>artificial sequence
<400> 22
ctcccgtctc tccacaacat 20
<210> 23
<211> 20
<212> DNA
<213>artificial sequence
<400> 23
tggtgtctgt aactgctgcc 20
<210> 24
<211> 20
<212> DNA
<213>artificial sequence
<400> 24
agttcttggg gaggaatcgt 20
<210> 25
<211> 20
<212> DNA
<213>artificial sequence
<400> 25
ttggtaaatc ggcatcttcc 20
<210> 26
<211> 20
<212> DNA
<213>artificial sequence
<400> 26
accgaaaagg gacaaggagt 20
<210> 27
<211> 20
<212> DNA
<213>artificial sequence
<400> 27
ggctcacttg tgcaacaaaa 20
<210> 28
<211> 20
<212> DNA
<213>artificial sequence
<400> 28
cgactgtcca cattggtgtc 20
<210> 29
<211> 20
<212> DNA
<213>artificial sequence
<400> 29
accttttcat gcaaattccg 20
<210> 30
<211> 20
<212> DNA
<213>artificial sequence
<400> 30
aggatcatgc attcggtttc 20
<210> 31
<211> 20
<212> DNA
<213>artificial sequence
<400> 31
gggtttgggt ttgagttgtg 20
<210> 32
<211> 20
<212> DNA
<213>artificial sequence
<400> 32
aaaagcaaac ggggctactt 20
<210> 33
<211> 20
<212> DNA
<213>artificial sequence
<400> 33
gtcgagagga aggagtggaa 20
<210> 34
<211> 20
<212> DNA
<213>artificial sequence
<400> 34
tcgttataat cgtccggctt 20
<210> 35
<211> 20
<212> DNA
<213>artificial sequence
<400> 35
ctccaccaca accacaacag 20
<210> 36
<211> 20
<212> DNA
<213>artificial sequence
<400> 36
ggttcacgtt ctctcgaagc 20
<210> 37
<211> 20
<212> DNA
<213>artificial sequence
<400> 37
gcttttaagg aaccgggaac 20
<210> 38
<211> 20
<212> DNA
<213>artificial sequence
<400> 38
agattcccac tgaatgtccg 20
<210> 39
<211> 22
<212> DNA
<213>artificial sequence
<400> 39
tttctgagat ttgacgagga aa 22
<210> 40
<211> 20
<212> DNA
<213>artificial sequence
<400> 40
actgcttgca ccgttgaact 20
<210> 41
<211> 20
<212> DNA
<213>artificial sequence
<400> 41
tgagaaaccc acaaagtccc 20
<210> 42
<211> 20
<212> DNA
<213>artificial sequence
<400> 42
gcgtcaaatc ccaatcactt 20
<210> 43
<211> 20
<212> DNA
<213>artificial sequence
<400> 43
tctagatctt tgagcgggga 20
<210> 44
<211> 20
<212> DNA
<213>artificial sequence
<400> 44
aatgccgtca atttctcacc 20
Claims (2)
1. wax-apple EST-SSR molecular labeling, it is characterised in that: the described wax-apple EST-SSR molecular labeling number be WASSR3,
WASSR12、WASSR13、WASSR37、WASSR38、 WASSR39、WASSR41、WASSR42、WASSR44、WASSR50、
WASSR55、WASSR60、WASSR63、WASSR70、WASSR72、WASSR74、WASSR80、WASSR85、WASSR87、
WASSR88, WASSR91 and WASSR92, the primer pair sequence of the corresponding wax-apple EST-SSR molecular labeling of each number is in order
As shown in SEQ ID NO.1-44.
2. wax-apple EST-SSR molecular labeling as described in claim 1 is in wax-apple analysis of genetic diversity and genetic map construction
In application.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710318661.2A CN106916901B (en) | 2017-05-08 | 2017-05-08 | Wax-apple EST-SSR molecular labeling |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710318661.2A CN106916901B (en) | 2017-05-08 | 2017-05-08 | Wax-apple EST-SSR molecular labeling |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106916901A CN106916901A (en) | 2017-07-04 |
| CN106916901B true CN106916901B (en) | 2019-08-27 |
Family
ID=59567914
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201710318661.2A Active CN106916901B (en) | 2017-05-08 | 2017-05-08 | Wax-apple EST-SSR molecular labeling |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN106916901B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN116516042B (en) * | 2023-05-18 | 2024-04-26 | 河北省农林科学院经济作物研究所 | SSR molecular marker related to fructus forsythiae germplasm resource identification, corresponding primer and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103233075A (en) * | 2013-05-09 | 2013-08-07 | 南京农业大学 | Method for developing dendranthema SSR (Simple Sequence Repeat) primer based on transcriptome sequencing |
| CN104673884A (en) * | 2014-05-24 | 2015-06-03 | 四川农业大学 | Method of developing polymorphic EST-SSR marker by utilizing complete genome and EST data |
| CN104846093A (en) * | 2015-05-14 | 2015-08-19 | 浙江大学 | Brassica juncea EST-SSR (expressed sequence tag-simple sequence repeat) marker primer group based on development of transcriptome sequence |
-
2017
- 2017-05-08 CN CN201710318661.2A patent/CN106916901B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103233075A (en) * | 2013-05-09 | 2013-08-07 | 南京农业大学 | Method for developing dendranthema SSR (Simple Sequence Repeat) primer based on transcriptome sequencing |
| CN104673884A (en) * | 2014-05-24 | 2015-06-03 | 四川农业大学 | Method of developing polymorphic EST-SSR marker by utilizing complete genome and EST data |
| CN104846093A (en) * | 2015-05-14 | 2015-08-19 | 浙江大学 | Brassica juncea EST-SSR (expressed sequence tag-simple sequence repeat) marker primer group based on development of transcriptome sequence |
Non-Patent Citations (1)
| Title |
|---|
| 莲雾转录组SSR信息分析及其分子标记开发;魏秀清等;《园艺学报》;20180331;第45卷(第3期);541-551 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN106916901A (en) | 2017-07-04 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103740711B (en) | Indel marker linked with yellow flesh gene yf of cucmis sativus L. and application of Indel marker | |
| CN113789408B (en) | Screening and application of SSR molecular marker primers for identifying variety of non-heading Chinese cabbage | |
| CN104928396A (en) | Method for rapidly identifying hot pepper species and golden pepper purity degree by using EST-SSR molecular markers | |
| CN106916901B (en) | Wax-apple EST-SSR molecular labeling | |
| CN102876790B (en) | Simple sequence repeat (SSR) marker polymerase chain reaction (PCR) reaction method applied to chrysanthemum and related species thereof universally | |
| CN108377901B (en) | Continuous multi-generation bud mutation seed selection and method for continuously improving apple red character by combining with crossbreeding | |
| CN106350591B (en) | Construct SSR primer, constructed finger-print and its application of coloured silk Ye Luanshu Variety fingerprinting | |
| CN113604598A (en) | Molecular marker primer and method for identifying common camellia oleifera and small camellia oleifera | |
| CN109652411A (en) | Fluorescent SSR primer combination and application thereof in construction of molecular fingerprint of new variety of white wax | |
| CN105543383A (en) | InDel site interlocked with red-peel/green-peel properties of pears as well as molecular marker based on site and application of molecular marker | |
| CN110063253A (en) | A kind of method of breeding high-yield shiny-leaved yellowhorn strain | |
| CN116144819B (en) | SNP molecular marker closely linked with main effect QTL of pumpkin pulp carotenoid and application of SNP molecular marker | |
| Andarini et al. | Genetic relationship of pigmented rice (Oryza sativa L.) collected from Eastern Indonesia based on morpho-agronomical traits and SSR markers | |
| CN112725521B (en) | Dendrobium chrysotoxum SSR molecular marker primer composition and application thereof | |
| CN106676175B (en) | For the Rapid identification oil tea breeding Cen specificity labeled primers pair and authentication method of soft No. 2 | |
| CN107815509A (en) | SSR primer sets and its application based on the exploitation of okra transcript profile sequence | |
| Guan et al. | Identification of superior clones by RAPD technology in Xanthoceras sorbifolia Bge. | |
| CN102827837A (en) | Cucumis sativus L. tender-fruit white-peel gene w linked SSR marker and use thereof | |
| CN108796111B (en) | Molecular marker primer for identifying kiwi fruit and golden plum varieties and application | |
| CN108642205A (en) | The method for developing jerusalem artichoke SSR molecular marker primer based on transcript profile sequence | |
| CN109280722A (en) | Musky gourd transcript profile SSR molecular marker primer and its application | |
| CN103820542A (en) | Method for identifying peach bud mutation | |
| CN109055599A (en) | Molecular labeling primer and application for the U.S. cultivar identification of Kiwi berry gold | |
| CN114438239B (en) | Molecular marker for identifying large 10 of mulberry variety Yuehong, identification primer group, kit and application | |
| CN109055600B (en) | Molecular marker primer for identifying variety No. 2 of kiwi fruit Hongtianhong and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20200527 Address after: 353199 No. 10, Jinggong Road, Jian'ou City, Nanping City, Fujian Province Patentee after: Fujian Chenxi Seed Industry Co., Ltd Address before: Jinan District of Fuzhou City, Fujian province 350013 Pu town Party Patentee before: POMOLOGY RESEARCH INSTITUTE, FUJIAN ACADEMY OF AGRICULTURAL SCIENCES |
|
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20210909 Address after: 332 Hankou village, Dapu Town, Yongchun County, Quanzhou City, Fujian Province Patentee after: Ye Shenghai Address before: 353199 No. 10 Jinggong Road, Jian'ou City, Nanping City, Fujian Province Patentee before: Fujian Chenxi Seed Industry Co.,Ltd. |
|
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20211014 Address after: Liu'an Development Zone, Yongchun County, Quanzhou City, Fujian Province (east side of Taoxi bridge) Patentee after: Yongchun County Product Quality Inspection Institute Fujian fragrance product quality inspection center, national incense burning product quality supervision and Inspection Center (Fujian) Address before: 332 Hankou village, Dapu Town, Yongchun County, Quanzhou City, Fujian Province Patentee before: Ye Shenghai |
|
| TR01 | Transfer of patent right |